The proposed studies are aimed at understanding the factors that regulate the development of human mast cells in general and, more specifically, differentiation and maturation of the two types of human mast cells known as MCT and MCTC. Human fetal livers and cord blood mononuclear cells will be used as sources of precursor cells. Maturation in vitro will be analyzed in a coculture system using murine 3T3 fibroblasts or in suspension cultures supplemented with various growth factors, including rhIL-3, rhIL-4 and rhSCF (stem cell factor). Cultured cells will be examined by immunohistochemistry for the presence of MCT and/or MCTC cells. Culture supernatants and extracts of cell pellets will be analyzed for their content of tryptase by an ELISA and of histamine by an RIA. Cellular content of chymase will be determined by an enzymatic assay. Electron microscopy of cultured cells will be performed at designated time points during the cultures. In situ hybridization will be used to detect tryptase mRNA in developing mast cells. These studies will be extended to encompass analysis of the type and state of differentiation of mast cells developing in vivo in fibrotic disorders, specifically scleroderma and keloid. These studies would improve our understanding of the potential role that one or the other type of mast cells may play in inflammatory conditions associated with fibrosis and, in turn, may lead to new therapeutic approaches for mast cell-related disorders.